摘要
本研究旨在优化红娘子蛋白提取工艺,考察红娘子生品及炮制品蛋白成分对肝癌细胞Bel-7402的影响。试验以蛋白含量为评价指标,考察料液比、超声时间、溶液pH对红娘子蛋白提取的影响,通过正交试验优选最佳提取工艺。体外研究红娘子生品及炮制品蛋白成分对Bel-7402肝癌细胞的影响,CellCountingKit-8检测对细胞活力影响;细胞集落研究对细胞增殖能力影响;划痕试验检测对细胞迁移能力影响;AnnexinV-FITC/PI双染法检测对细胞凋亡影响;WesternBlotting检测相关凋亡蛋白的作用机制。红娘子蛋白最佳提取工艺为红娘子粉末加20倍体积水溶液(pH为10.8),冰水浴超声提取150min。生品及炮制品蛋白可显著降低Bel-7402肝癌细胞存活率,具有良好的抑制肝癌细胞增殖能力,可显著抑制肝癌细胞迁移;显著增加Bel-7402肝癌细胞凋亡率,可促进凋亡相关蛋白BCL2-AssociatedX(Bax)和Caspase-3表达,降低B-cell lymphoma-2(Bcl-2)蛋白表达。优化的红娘子蛋白提取工艺稳定,蛋白含量可达240.55mg/g。红娘子生品及炮制品蛋白成分均具有较好的抑制Bel-7402肝癌细胞迁移及增殖的能力,其机制可能是通过上调抗凋亡蛋白Bcl-2蛋白表达、下调促凋亡蛋白Bax蛋白表达,激活Caspase级联反应,上调效应子Caspase-3蛋白表达,从而促进细胞凋亡,且炮制品蛋白效果优于生品蛋白。
The extraction process of Huechys sanguinea(De Geer)protein was optimized and the effect of the protein components of the raw and processed H.sanguinea on the liver cancer cells Bel-7402 was investigated.Taking the protein content as the evaluation index,the effects of solid-liquid ratio,ultrasonic time and solution pH on the extraction of H.sanguinea protein were investigated,and the optimal ex-traction process was optimized by orthogonal experiment.The effects the protein components of the raw and processed H.sanguinea on Bel-7402 hepatoma cells were studied in vitro.The effect of Cell counting Kit-8 detection on cell viability was detected;The effect of cell colony study on cell proliferation and scratch test was used to detect the effect on cell migration;Annexin V-FITC/PI double staining was used to detect the effect on apoptosis.Western blotting was used to detect the mechanism of apoptosis of related proteins.The optimum ex-traction process ofH.sanguinea protein was as follows:red damselfly powder plus 20 times volume of water solution(pH is 10.8),and ultra-sonic extraction in ice water bath for 150 min.Raw and processed protein can significantly reduce the survival rate of Bel-7402 hepatoma cells,which have a good ability to inhibit the proliferation of liver cancer cells,significantly inhibit the migration of liver cancer cells and the apoptosis rate of Bel-7402 hepatoma cells was significantly increased.It can promote the expression of apoptosis related proteins BCL2 Associated X(Bax)and Caspase-3,and reduce the expression of B-cell lymphoma-2(Bcl-2)protein.The optimized extraction process of H.sanguinea protein is stable,and the protein content can reach 240.55 mg/g.The protein components of raw and processed products of H.sanguinea have a better ability to inhibit the migration and proliferation of by increasing the expression of anti-apoptotic protein Bcl-2 and decreasing the expression of pro-apoptotic protein Bax,the caspase cascade reaction is activated and the expression of effector Caspase-3 protein is increased,thus promoting cell apoptosis.The effect of processed protein is better than that of raw protein.
作者
姜蓉
李正龙
王淑敏
JIANG Rong;LI Zhengong;WANG Shumin(College of Pharmacy,Changchun University of Traditional Chinese Medicine,Changchun 130117,China)
出处
《特产研究》
2025年第2期73-80,共8页
Special Wild Economic Animal and Plant Research
基金
吉林省中药材标准及中药饮片炮制规范项目(JLPZGF-2020-082)。
作者简介
共同第一作者:姜蓉(1995-),女,安徽滁州人,硕士,从事中药健康产品研究与开发;共同第一作者:李正龙(1997-),男,安徽阜阳人,硕士,从事中药健康产品研究与开发;通讯作者:王淑敏(1965-),女,吉林长春人,博士,教授,博士研究生导师,从事中药炮制关键技术及科学内涵研究。
